The microenvironment of a tissue or organ has a profound impact on the development and manifestations of local immune responses. For example, parenchymal cells and blood vessels define compartments in which lymphocytes and antigen presenting cells (APC) interact and generate metabolites such as nitric oxide (NO) and cytokines that modulate lymphocyte responses. As another example, cell adhesion molecules expressed on endothelial cell surfaces control the egress, activation and behavior of lymphocytes. Of the several components of a tissue or organ microenvironment that might influence immune responses, this application focuses on heparan sulfate proteoglycan, a biologically active protein- polysaccharide conjugate that regulates cell interactions in a number of systems. The overall objective of the research is to evaluate the contribution of the microenvironment of an allograft to the development and manifestations of allograft rejection focusing in particular on the metabolism of endothelial cell associated heparan sulfate proteoglycan. The first aim is to elucidate mechanisms responsible for alteration of heparan sulfate metabolism in an organ allograft. This aim will be addressed by characterizing an enzyme system responsible for causing release of heparan sulfate from endothelial cells and exploring the consequences of interrupting that system in a murine renal allograft model. The second aim is to determine the physiologic implications of the release of heparan sulfate from endothelial cells and exposure of endothelial cells to soluble heparan sulfate. This aim will be pursued by testing the physiologic properties of endothelial cells following cleavage and release of heparan sulfate from the cells and following exposure of the cells to heparan sulfate. The third aim will involve development of novel strategies to inhibit heparan sulfate release in allogeneic organ grafts. These strategies will be tested for their ability to prevent the untoward metabolism of heparan sulfate proteoglycan and for the effects on the outcome of experimental renal allografts.